The ability to precisely monitor the effectiveness of therapy for non-Hodgkin lymphoma has important clinical implications. In patients with curable lymphomas, such as diffuse large B-cell lymphoma, the eradication of all disease is necessary for cure. In patients with incurable lymphomas, such as follicular lymphoma and mantle cell lymphoma, deep and durable remissions are associated with improvements in survival. Radiographic imaging modalities such as computed tomography and positron emission tomography are the current gold standard for monitoring therapy, but they are fundamentally limited by radiation risks, costs, lack of tumor specificity, and inability to detect disease at the molecular level. Novel sequencing-based methods can detect circulating tumor DNA (ctDNA) in the peripheral blood with great sensitivity, which opens new opportunities for molecular monitoring before, during, and after therapy. Beyond monitoring, ctDNA can also be used as a “liquid biopsy” to assess for molecular changes after therapy that may identify treatment-resistant clones. ctDNA is an emerging tool that may transform our ability to offer precision therapy in non-Hodgkin lymphoma.
Non-Hodgkin lymphomas are heterogeneous lymphoid malignancies that are managed with a range of therapeutic goals. Aggressive B-cell lymphomas such as diffuse large B-cell lymphoma (DLBCL) are typically treated with curative intent, and frontline therapy can achieve a cure in up to 70% of patients. In the current rituximab era, however, patients with refractory DLBCL or those who relapse after therapy are not commonly cured with standard approaches such as autologous stem cell transplant. Novel therapies are emerging for DLBCL that target mechanisms of cellular resistance, but they take years to develop and benefit only subsets of patients.
Incurable lymphomas, such as follicular lymphoma (FL) and mantle cell lymphoma (MCL), exhibit a perpetually relapsing disease course, and the primary goal of therapy is induction of deep and durable remissions.[4,5] Increasingly, patients with these lymphomas are treated for extended periods with maintenance therapies. Clinical decisions regarding the duration of maintenance therapy for indolent lymphomas are usually made empirically and without direct measurement of disease burden or molecular aberrations within the tumor.
The gold standard for initial staging, response assessment, and surveillance monitoring in non-Hodgkin lymphomas is assessment with imaging modalities such as computed tomography (CT) and fluorodeoxyglucose (FDG) positron emission tomography (PET). Despite widespread use, imaging scans have important drawbacks, including radiation exposure, cost, and low tumor specificity.[7-10] In addition, PET scans do not perform well as surveillance tools, and there is concern regarding the use of imaging for detection of asymptomatic relapse.[11,12] PET scans have also been extensively tested as interim biomarkers for purposes of tailoring therapy. Despite initial enthusiasm, two large studies failed to demonstrate a benefit to switching therapy in patients with DLBCL on the basis of interim PET (iPET) results.[13,14]
Precision medicine approaches to non-Hodgkin lymphomas consider the molecular profile of the tumor when targeted therapies are selected. Currently, the standard method for determining tumor genetic profiles requires tissue biopsies. Tissue biopsies, however, contain procedural risks, are subject to sampling error, and cannot account for spatial tumor heterogeneity. In view of these limitations, genetic profiles obtained from peripheral blood are of great interest.
Novel sequencing-based assays can detect cell-free circulating tumor DNA (ctDNA), which encodes the immunoglobulin gene sequence unique to individual B-cell lymphomas.[16-20] Assays involving ctDNA that detect somatic mutational profiles and that can serve as a “liquid biopsy” to monitor for the development of treatment-resistant clones are also being developed for non-Hodgkin lymphomas. Both patients with indolent non-Hodgkin lymphomas and those with aggressive non-Hodgkin lymphomas stand to benefit from advances in technologies that more precisely define disease relapse at earlier time points. Precision molecular monitoring of ctDNA offers lower detection limits than scans and may overcome the limitations of both imaging scans and tissue biopsies (Figure 1). In this review, we discuss the potential applications of monitoring ctDNA in patients with DLBCL, FL, and MCL.
Advances in Molecular Monitoring Methods for Non-Hodgkin Lymphomas
The quest to find tumor-specific molecules in the peripheral blood to monitor patients with non-Hodgkin lymphoma dates back decades. The principal barriers to widespread use have been difficulties in identifying appropriate targets with broad clinical applicability across all non-Hodgkin lymphoma subtypes. In specific subtypes, recurrent translocations can be detected in the blood by polymerase chain reaction (PCR)-based methods (Figure 2). These include the IGH-CCND1 translocation, t(11;14), in MCL and the IGH-BCL2 translocation, t(14;18), in FL. All patients with MCL and the majority of patients with FL harbor these respective translocations, which makes them attractive targets for molecular monitoring.[22,23] Allele-specific oligonucleotide PCR (ASO-PCR) techniques use patient-specific primers to detect and quantitate BCL2 and CCND1 rearrangements, with a limit of detection approaching 1 × 105 cells. Because of variations in breakpoint regions, however, PCR is not able to detect IGH-BCL2 translocations in the blood of all patients with FL. Furthermore, most non-Hodgkin lymphoma subtypes do not have recurrent translocations amenable to molecular monitoring. Given the need for patient-specific primers and the lack of universal applicability, ASO-PCR is considered laborious and not conducive to widespread use.
Tumor-specific DNA fragments can be detected noninvasively in the blood in both cellular (circulating tumor cells [CTCs]) and cell-free (ctDNA) forms (see Figure 2). In certain lymphomas, such as DLBCL, however, CTCs are rare, and recent data indicate a possible relationship between defective homing pathways and systemic dissemination.[24-26] Multiple studies have demonstrated higher levels of circulating cell-free DNA (cfDNA) in cancer patients than in healthy controls, which result from the constant shedding of DNA fragments into the peripheral blood from cancer cells undergoing apoptosis, secretion, and necrosis.[27-29] Since cfDNA can originate from both malignant and nonmalignant sources, assays for ctDNA must accurately discriminate the tumor-specific DNA fragments within the overall cfDNA concentration. ctDNA is therefore defined as the tumor-specific DNA sequences found in either the plasma or the serum of the blood and may represent as little as 0.01% of total cfDNA. An early study using real-time quantitative reverse transcriptase-PCR of the germline beta-globin gene revealed significantly higher median cfDNA levels in patients with DLBCL, MCL, and Hodgkin lymphoma than in healthy controls, with similar concentrations in patients with FL. This study was the first to validate ctDNA as a quantifiable tumor-specific biomarker in patients with non-Hodgkin lymphoma.
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